A novel mechanism underlying allosteric regulation of ADAMTS-13 revealed by hydrogen-deuterium exchange plus mass spectrometry.
Res Pract Thromb Haemost
; 7(1): 100012, 2023 Jan.
Article
em En
| MEDLINE
| ID: mdl-36852110
ABSTRACT
Background:
ADAMTS-13, a plasma metalloprotease, cleaves von Willebrand factor. ADAMTS-13 activity appears to be regulated through allosteric inhibition by its distal C-terminus.Objectives:
The objective of this study was to better understand how domain-domain interactions may affect ADAMTS-13 conformations and functions.Methods:
We performed deuterium-hydrogen exchange plus mass spectrometry to assess the number and rate of deuterium incorporation into various peptides of full-length ADAMTS-13 and its truncated variants.Results:
Under physiological conditions, a bimodal distribution of deuterium incorporation was detected in the peptides from metalloprotease (217-230 and 282-304), cysteine-rich (446-482), and CUB (for complement C1r/C1s, Uegf, Bmp1) domains (1185-1214, 1313-1330, 1341-1347, 1358-1378, and 1393-1407) of full-length recombinant ADAMTS-13, but not of truncated variants. These results suggest that the full-length ADAMTS-13 undergoes conformational changes. On removal of the middle and distal C-terminal domains, the number and rate of deuterium incorporation were increased in the peptides from cysteine-rich (445-467, 467-482, and 495-503) and spacer domains (621-642 and 655-654) but decreased in the peptides from metalloprotease (115-124, 217-230, and 274-281). Moreover, most peptides, except for 217-230 and 1357-1376, exhibited a pD-dependent deuterium incorporation in the full-length ADAMTS-13, but not in the truncated variant (eg, MDTCS or T5C). These results further suggest that the bimodal deuterium incorporation observed in the peptides from the full-length ADAMTS-13 is the result of potential impact from the middle to distal C-terminal domains. Surface plasmon resonance revealed the direct binding interactions between the distal and proximal domains of ADAMTS-13.Conclusion:
Our results provide novel insight on how intramolecular interactions may affect conformations of ADAMTS-13, thus regulating its proteolytic functions.
ADAMTS-13 A disintegrin and metalloprotease with thrombospondin type 1 repeats-13, MDTCS A C-terminal truncated ADAMTS-13 variant after the spacer domain; ADAMTS-13 protein; DTCS An ADAMTS-13 variant consisting of a disintegrin, first TSP1 repeat, Cys-rich, and spacer domain., TTP Thrombotic thrombocytopenic purpura; HX-MS hydrogendeuterium exchange plus mass spectrometry, ULVWF Ultra large von Willebrand factor; T5C A C-terminal fragment of ADAMTS-13 consisting of thrombospondin type 1 repeat 5-8 and CUB domain., CUB The CUB domain for complement C1r/C1s, Uegf, Bmp1; allosteric regulation; conformation; deuterium exchange
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Idioma:
En
Revista:
Res Pract Thromb Haemost
Ano de publicação:
2023
Tipo de documento:
Article
País de afiliação:
Estados Unidos
País de publicação:
EEUU
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ESTADOS UNIDOS
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ESTADOS UNIDOS DA AMERICA
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EUA
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UNITED STATES
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UNITED STATES OF AMERICA
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US
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USA